In Vitro Refolding of Triosephosphate Isomerase from <Emphasis Type="Italic">L. donovani</Emphasis> |
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Authors: | Kishore Kumar Prachi Bhargava Uma Roy |
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Institution: | (1) Division of Biochemistry, Central Drug Research Institute, Lucknow, 226001, UP, India; |
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Abstract: | The triosephosphate isomerase of Leishmania donovani (LdTIM) was expressed at high level in Escherichia coli. The TIM gene was cloned in expression vector pET-23(a) with C-terminal 6× His tag fused in frame, and expressed as a 27.6-kDa
protein in E. coli as inclusion bodies. The recombinant LdTIM from E. coli lysate was solubilized in 6 M guanidine hydrochloride and purified by Ni-NTA chromatography. In the present study, the effect
of bovine serum albumin on the reactivation of TIM was investigated. Furthermore, 8-anilino-1-naphthalene sulfonic acid was
used to detect the structural changes induced by bovine serum albumin (BSA). Here, we conclude that BSA assists in the refolding
and regain of LdTIM enzyme activity by providing framework for structure formation. This study indicates that numerous protein–protein
contacts are constantly occurring inside the cell that leads to the formation of native protein. |
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